The ultimate goal of our research project is to determine the genetic and molecular basis of peptide hormone's cell surface receptor-mediated signal transfer mechanisms. The genetic variations in the regulatory receptor molecules, the nature of hormonal signals, the sequence of events in hormone-induced biological responses are being studied by the strategy of mutagenesis and somatic cell genetics in combination with biochemical techniques. Two bioactive polypeptides have been chosen, insulin and epidermal growth factor (EGF). These hormones are involved in the control of cellular proliferation and have served as basic model systems for the study of physiology and dynamics of ligand/receptor interaction, membrane modulation and malignant transformation. A series of insulin (or EGF)-nonresponsive variants are being selected from in vitro mutagenized cell population of established cell lines, such as Swiss/3T3. A new selection procedure has been devised which utilizes the cytotoxic hybrid molecules crosslinked by disulfide bond between insulin (or EGF) and diphtheria toxin fragment A (or ricin A subunit). In combination of other selection agents, it is expected that various states of deficiency will be found in receptor and post-receptor functions. The mutants are being characterized both genetically and biochemically. The mutants of mouse origin are fused with human cells in order to perform mapping of human genes which are responsible for production and regulation of functional receptors and other components.